Recent advances in tumor biology have led to the identification of a variety of intracellular oncogenic pathways as potential targets for cancer therapy. Specifically, many studies have found that activation of the JAK/STAT pathway promotes tumor cell proliferation and survival in various types of leukemia and lymphoma. Our preliminary data demonstrate aberrantly activated JAK2 and STAT3 in more than 50% of diffuse large B-cell lymphoma patient samples. In vitro inhibition of JAK2 with the novel JAK2 inhibitor TG101348 (TG) inhibited JAK2 and STAT3 phosphorylation and induced apoptosis in a variety of lymphoma cell lines and patient samples. In this proposal the overall goal is to identify the molecular mechanisms underlying activation ofthe JAK/STAT pathway in lymphoma and to learn if inhibitors of this pathway can produce clinical benefit. We have identified several novel missense mutations in JAK2 and STAT3 genes. In Aim 1 we will characterize the biological and therapeutic significance of these mutations with a site-directed mutagenesis approach. Suppressors of cytokine signaling (S0CS1) and protein tyrosine phosphatases (SHP1) are known key negative regulators of the JAK/STAT pathway. Our preliminary data demonstrate silencing of SHP1 and S0CS1 genes in 33% and 86%, respectively, of DLBCL lymphoma samples. In Aim 2, we will delineate the mechanisms of silencing and how this regulates JAK/STAT pathway activation. The JAK/STAT signaling pathway is utilized by a number of growth factors and cytokines. We have identified increases in several JAK/STAT pathway-specific cytokines (IL-2, IL-6, IL-10 and EGF) in serum samples from patients with lymphoma compared to normal controls. In vitro we found in lymphoma cells that JAK2 and STAT3 are rapidly activated in response to IL-10. Aim 3 will investigate the role of signaling for these interieukins mediated through their receptors with a focus on IL-10. This project is based on solid preliminary data demonstrating that the JAK/STAT pathway is a key mechanism for lymphoma growth and survival. These data have guided the design ofthe phase II trial in Aim 4 that will test TG in patients with relapsed lymphoma. Correlative research using patient samples pre- and posttherapy with JAK/STAT pathway inhibitor will increase our understanding ofthe mechanisms of how this pathway is regulated at the molecular and genetic level. These basic and clinical studies, working together, aim to offer a new therapeutic approach for patients with lymphoma.